Severe drought exposure in utero associates to children’s epigenetic age acceleration in a global climate change hot spot

The goal of this study is to examine the association between in utero drought exposure and epigenetic age acceleration (EAA) in a global climate change hot spot. Calculations of EAA in adults using DNA methylation have been found to accurately predict chronic disease and longevity. However, fewer studies have examined EAA in children, and drought exposure in utero has not been investigated. Additionally, studies of EAA in low-income countries with diverse populations are rare. We assess EAA using epigenetic clocks and two DNAm-based pace-of-aging measurements from whole saliva samples in 104 drought-exposed children and 109 same-sex sibling controls in northern Kenya. We find a positive association between in utero drought exposure and EAA in two epigenetic clocks (Hannum’s and GrimAge) and a negative association in the DNAm based telomere length (DNAmTL) clock. The combined impact of drought’s multiple deleterious stressors may reduce overall life expectancy through accelerated epigenetic aging.


2024-4-16
• Epigenetic clocks were derived using R package methylCIPHER from MorganLevineLab, R package dnaMethyAge from Github, R function derived from original clock paper, and R code provided by by the authors of of "DNA methylation GrimAge version 2" 2" • Cell type proportions were estimated using R package EpiDISH.
• Figures 1 and 3 were generated using Microsoft Excel for Mac Version 16.83.
• Figure 4 was generated using R packages ggcorrplot and patchwork.
• All code is is run in in R version 4.1.2• Statistical analyses were conducted using the following packages: MCMCglmm, lmerTest and lme4.
• R code for data analyses is is available at at https://github.com/DuyNgoStats/BayesianEAA.The study's used biological sex assigned at birth as reported by parents and recorded in vaccination records, and consistent with Samburu cultural categories.The study design is maternal and same-sex sibling pair design (e.g., control siblings matched drought-exposed children; some families had more than one control sibling).All mothers identified as female based on sex assigned at birth and were the biological mothers of the children.Mothers (female) N = 104; Children, male = 98; female = 115.Total Child N = 213.Children's ages (1.81 -9.61 years) were documented with vaccination records and birth certificates and calculated to age in years (minimum of two decimal points).Sex of all participating children as identified by parents and children matched sex reported in health records.
The study recruited Samburu pastoralists, all of whom self-identify as Samburu ethnicity, East Africans in northern Kenya.
Samburu pastoralists are subsistence livestock herders living primarily in Samburu County, with some also living in adjacent Laikipia County.They engage in a climate change vulnerable livelihood, raising cows, goats, sheep, and, in some families, camels.Additionally, Samburu County is in a global hotspot for climate change vulnerability.Many Samburu families practice polygyny, with up to 5 wives.Gender inequality is substantial, with husbands controlling wives' labor and access to key elements of subsistence (e.g., access to livestock to milk).Women experience high rates of interpersonal spousal violence, which was documented in the overall study.
Samburu live in dispersed homesteads across a rugged landscape, with many homesteads accessible only off road or on foot.The drought catchment area consisted of all rural communities within multiple subregions of Samburu County in which drought effects on livestock mortality were documented by drought monitoring agencies, including lower rainfall areas of the Leroghi Plateau.Recruitment efforts extended to both remote and more accessible communities within those subregions to avoid proximity bias.Recruitment occurred through off road 4WD vehicle and motorcycle transport, reaching all communities within the catchment area at meetings advertised in advance by local village administrators.123 families were initially recruited, of whom, 11 were found not to meet inclusion criteria at recruitment and 1 family met most criteria but the mother had spent the majority of the pregnancy in Nairobi.Of the 111 families meeting all criteria, 4 families declined to participate, and 3 families agreed but were unavailable during the measurement and biospecimen visit due to long distance herding to follow pasture.No self-selection bias was evident, as all families with individuals who had been pregnant during the drought attended the meetings and were recruited.The number of families declining (4) was 3.6% and did not differ by any known socioeconomic or other characteristics.

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Describe how sample size was determined, detailing any statistical methods used to predetermine sample size OR if no sample-size calculation was performed, describe how sample sizes were chosen and provide a rationale for why these sample sizes are sufficient.
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The study employed mixed quantitative and qualitative methods, utilizing a same-sex sibling group design (exposed/unexposed in utero) to collect data based on the natural observational experiment of a severe drought occurring in 2008-2009 in northern Kenya.
Samburu pastoralist families exposed to the drought were recruited.The sample is comprised of 104 mothers exposed to the drought during pregnancy, their 104 live born offspring of those pregnancies, and 109 offspring of those mothers who are the samesex siblings of drought-exposed offspring (child ages 1.81 -9.61 years).The sample is representative: all families meeting the criteria in the catchment area were included and the number of families declining was small (94% of 111 families meeting criteria participated, with only 4 declining & 3 unavailable -6%).The sample was selected based on a rigorous same-sex sibling design, and the community selected based on authors' established working rapport and trust with the Indigenous community and their established exposure to severe drought.
The study relied on purposive sampling, recruiting throughout rural areas of the dispersed, semi-nomadic target county affected by the 2008-2009 severe drought.All individuals who met the criteria for the same-sex sibling design who agreed to participate were enrolled.The target was a minimum of 100 sibling pairs, based on similar studies (Cao-Lei, Lei, Renaud Massart, Matthew J. Suderman, Ziv Machnes, Guillaume Elgbeili, David P. Laplante, Moshe Szyf, Suzanne King.Study team members at the field site consisted of two senior data collection team members, 4 multilingual Samburu research assistants, and up to 3 Kenyan and U.S. undergraduate and graduate students at a time.Team members visited Samburu study partners at locations convenient to study partners (usually their homes).To enhance accuracy and rapport, team members were assigned tasks that they exclusively performed (e.g., anthropometric measurements, ethnographic interviews with mothers, behavioral observations, etc.)All observations and interviews were recorded with pen and paper, with paired team members to cross-check each other's work during data collection.Saliva samples for DNA methylation data were obtained using Oragene 600 kits.Researchers were not blind to hypotheses.
Rolling recruitment was initiated in October, 2017 and concluded in December 2018.Data collection occurred alongside recruitment and concluded in July 2019.
No data were excluded from analyses.4 families declined.Consistent with ethical protocol, reasons were not obtained.3 families were unavailable during biospecimen collection due to following their herds.Data was collected from 94% of eligible families.
In this natural experiment study, children exposed to drought = the experimental group and their same-sex unexposed siblings = control group.Note the sampling procedure.Describe the statistical methods that were used to predetermine sample size OR if no sample-size calculation was performed, describe how sample sizes were chosen and provide a rationale for why these sample sizes are sufficient.
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Western Michigan University for HSIRB; collaborative with University of Nairobi.Site approval was obtained at the Samburu County and local community level, and Samburu community leaders and study partners were collaboratively engaged at all phases of the study.The study was initiated prior to 2018 IRB rules and is subject to reconsent based on individual subject and Indigenous community agreements.
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